The newly defined RhoH gene has been demonstrated to be mutated in lymphoma samples. These alterations include chromosomal rearrangements and a high frequency of somatic mutations (up to 46 percent) in non-Hodgkin's lymphomas and diffuse large B-cell lymphoma. The RhoH gene encodes a novel hematopoietic-specific member of the RhoE subfamily, which is GTPase-deficient. Therefore RhoH does not cycle between GDP-bound and GTP-bound states and remains permanently in the active state. Thus the activity of RhoH is likely directly related to the level of the protein expressed in the cell. Previous studies have not directly measured RhoH expression in hematopoietic tumors. The biological role of RhoH in normal and malignant hematopoietic cell development and function have also not been studied. We hypothesize that expression level of RhoH is critical for its biological function in normal hematopoiesis and dysregulated expression may contribute to the transformed phenotypes in human lymphomas. Preliminary data support this hypothesis, since overexpression of RhoH results in abnormal proliferation and actin-based function of primary hematopoietic cells. In this grant, we propose to use a mouse model to elucidate the role and mechanism of RhoH in controlling growth, differentiation, transformation and related signaling pathways in hematopoietic cells, and examine functional interactions between RhoH, Rac and downstream effectors, group A Paks. In addition to this genetic approach, we will assess RhoH expression levels and their correlation with somatic mutations in the human RhoH gene and clinical phenotypes in diffuse large-B cell lymphomas. Specific Aim 1 will determine the role of RhoH in normal and malignant hematopoietic cells. In Specific Aim 2, we will determine RhoH-mediated signaling pathways that are critical in regulation of hematopoietic cell proliferation and function. In Specific Aim 3, we will examine the essential function of RhoH and its interaction with Rac using RhoH-deficient and Rac-deficient compound mice. These biological experiments may provide mechanistic insights and biological context to the previous findings of RhoH as a hypermutable gene in human cancer and possibly establish an animal model for Rho GTPases in tumorigenesis. Given the lineage-restricted expression of RhoH such an observation could also have important therapeutic implication.